A acne é uma condição da pele multifatorial com implicações socioeconômicas importantes. Um dos principais fatores que contribuem com a sua etiologia é a superprodução de sebo. Até o momento, há poucos tratamentos seguros e eficazes disponíveis. O receptor de melanocortina-5 (MC5R), um receptor acoplado à proteína G da família das rodopsinas, é uma das proteínas responsáveis pela diferenciação de sebócitos e consequente produção de sebo, mas não há opções de tratamento através do antagonismo deste receptor. Neste trabalho, investigamos a melanocortina-5 como alvo molecular para a descoberta de ligantes como moduladores da sebogênese. Para tanto, empregamos estudos de modelagem por homologia e triagem virtual baseada em estrutura do alvo para construir um modelo 3D da MC5R e identificar de candidatos a ligantes da proteína, respectivamente. Em seguida, avaliamos o potencial de inibição da sebogênese em sebócitos SEBO662AR em meio lipogênico. Os resultados obtidos indicaram a descoberta de peptídeos e flavonoides com características inibidoras e estimuladoras da produção de sebo. Novos esqueletos moleculares foram identificados como promissores para a modulação da sebogênese. Os estudos realizados permitirão o desenvolvimento de novos ativos dermatológicos e cosméticos com potencial de modular a oleosidade da pele, de modo a contribuir com a mitigação dos efeitos da acne, psoríase, alopecia e seborreia, entre outras doenças. / Acne is a multifactorial skin condition with important socioeconomic implications. One of the main factors that contribute with its etiology is sebum overproduction. Until now, there are few safe, effective treatments available. Melanocortin-5 receptor (MC5R), a G protein-coupled receptor of the rhodopsin family, is one of the proteins responsible for sebocyte differentiation and consequent sebum production, but there are no options for treatment by antagonism of this receptor. In this work, we investigated MC5R as molecular target for the discovery of ligands as sebogenesis modulators. For that, we used homology modeling studies, and structure-based virtual screening in order to, respectively, build a MC5R 3D model and identify ligand candidates for this protein. Then, we evaluated their sebogenesis inhibition potential on SEBO662AR sebocytes in lipogenic conditions. The obtained results indicated the discovery of peptides and flavonoids with inhibitory and stimulatory sebum production characteristics. New scaffolds were identified as promising for sebogenesis modulation. The performed studies will allow the development of novel dermatologic and cosmetic actives with the potential to modulate skin oiliness in order to contribute to the mitigation of the effects of acne, psoriasis, alopecia and seborrhea, among other diseases.

alpha-melanocyte stimulating hormone (alpha-MSH) is an important regulator of food intake, metabolic rate, and inflammation. Recently, alpha-MSH was shown to influence sympathetic activity and blood pressure regulation. In the present study, we investigated the cardiovascular effects of alpha-MSH in the nucleus tractus solitarii (NTS) of spontaneously hypertensive rats (SHR). Because nitric oxide (NO) is well-known to involve in central cardiovascular regulation, we elucidated the role of NO in the cardiovascular responses induced by alpha-MSH. In urethane-anesthetized SHR, unilateral microinjection of alpha-MSH (0.3-300 pmol) into the NTS produced dose-responsive depressor and bradycardic effects. The cardiovascular effects of alpha-MSH were abrogated by the antagonist of melanocortin receptor (MC3/4-R), SHU9119. Pretreatment with precursor of nitric oxide, L-arginine, enhanced the duration of alpha-MSH-mediated hypotensive effects, whereas prior application of L-NAME, a universal inhibitor of nitric oxide synthase (NOS), significantly attenuated the effects of alpha-MSH. Prior injection with inhibitor of inducible NOS, aminoguanidine, but not inhibitor of neuronal NOS, 7-nitroindazole, attenuated the hypotensive effect of alpha-MSH. In summary, these results indicated alpha-MSH induced depressor and bradycardic effects in the NTS of SHR. Besides, the hypotensive mechanism of alpha-MSH was mediated via MC4-R and involved with iNOS activation in the NTS of SHR.

Various molecular pathways in the brain shape our understanding of good and bad, as well as our motivation to seek and avoid such stimuli. This work evolves around how systemic inflammation causes aversion; and why general unpleasant states such as sickness, stress, pain and nausea are encoded by our brain as undesirable; and contrary to these questions, how drugs of abuse can subjugate the motivational neurocircuitry of the brain. A common feature of these various disease states is involvement of the motivational neurocircuitry - from mesolimbic to striatonigral pathways. Having an intact motivational system is what helps us evade negative outcomes and approach natural positive reinforcers, which is essential for our survival. During disease-states the motivational neurocircuitry may be overthrown by the molecular mechanisms that originally were meant to aid us. In study I, to investigate how inflammation is perceived as aversive, we used a behavioral test based on Pavlovian place conditioning with the aversive inflammatory stimulus E. coli lipopolysaccharide (LPS). Using a combination of cell-type specific gene deletions, pharmacology, and chemogenetics, we uncovered that systemic inflammation triggered aversion by MyD88-dependent activation of the brain endothelium followed by COX1-mediated cerebral prostaglandin E2 (PGE2) synthesis. Moreover, we showed that inflammation-induced PGE2 targeted EP1 receptors on striatal dopamine D1 receptor–expressing neurons and that this signaling sequence induced aversion through GABA-mediated inhibition of dopaminergic cells. Finally, inflammation-induced aversion was not an indirect consequence of fever or anorexia but constituted an independent inflammatory symptom triggered by a unique molecular mechanism. Collectively, these findings demonstrate that PGE2-mediated modulation of the dopaminergic circuitry is a key mechanism underlying inflammation-induced aversion. In study II, we investigate the role of peripheral IFN-γ in LPS induced conditioned place aversion by employing a strategy based on global and cell-type specific gene deletions, combined with measures of gene-expression. LPS induced IFN-ɣ expression in the blood, and deletion of IFN-ɣ or its receptor prevented conditioned place aversion (CPA) to LPS. LPS increased the expression of chemokine Cxcl10 in the striatum of normal mice. This induction was absent in mice lacking IFN-ɣ receptors or Myd88 in blood brain barrier endothelial cells. Furthermore, inflammation-induced aversion was blocked in mice lacking Cxcl10 or its receptor Cxcr3. Finally, mice with a selective deletion of the IFN-ɣ receptor in brain endothelial cells did not develop inflammation-induced aversion. Collectively, these findings demonstrate that circulating IFN-ɣ binding to receptors on brain endothelial cells which induces Cxcl10, is a central link in the signaling chain eliciting inflammation-induced aversion. In study III, we explored the role of melanocortin 4 receptors (MC4Rs) in aversive processing using genetically modified mice in CPA to various stimuli. In normal mice, robust aversions were induced by systemic inflammation, nausea, pain and kappa opioid receptor-induced dysphoria. In sharp contrast, mice lacking MC4Rs displayed preference towards most of the aversive stimuli, but were indifferent to pain. The unusual flip from aversion to reward in mice lacking MC4Rs was dopamine-dependent and associated with a change from decreased to increased activity of the dopamine system. The responses to aversive stimuli were normalized when MC4Rs were re-expressed on dopamine D1 receptor-expressing cells or in the striatum of mice otherwise lacking MC4Rs. Furthermore, activation of arcuate nucleus proopiomelanocortin neurons projecting to the ventral striatum increased the activity of striatal neurons in a MC4R-dependent manner and elicited aversion. Our findings demonstrate that melanocortin signaling through striatal MC4Rs is critical for assigning negative motivational valence to harmful stimuli. The neurotransmitter acetylcholine has been implied in reward learning and drug addiction. However, the role of cholinergic receptor subtypes in such processes remains elusive. In study IV we investigated the function of muscarinic M4Rs on dopamine D1R expressing neurons and acetylcholinergic neurons, using transgenic mice in various reward-enforced behaviors and in a “waiting”-impulsivity test. Mice lacking M4-receptors from D1-receptor expressing neurons exhibited an escalated reward seeking phenotype towards cocaine and natural reward, in Pavlovian conditioning and an operant self-administration task, respectively. In addition, the M4-D1RCre mice showed impaired waiting impulsivity in the 5-choice-serial-reaction-time-task. On the contrary, mice without M4Rs in acetylcholinergic neurons were unable to learn positive reinforcement to natural reward and cocaine, in an operant runway paradigm and in Pavlovian conditioning.  Immediate early gene expression mirrored the behavioral findings arising from M4R-D1R knockout, as cocaine induced cFos and FosB was significantly increased in the forebrain of M4-D1RCre mice, whereas it remained normal in the M4R-ChatCre mice. Our study illustrates that muscarinic M4Rs on specific neural populations, either cholinergic or D1R-expressing, are pivotal for learning processes related to both natural reward and drugs of abuse, with opposing functionality.